Seasonal Changes of Circulating Sex Steroid Hormones Associated With Vomeronasal Function of the Male Muskrats ( <i>Ondatra zibethicus</i> )

The vomeronasal organ (VNO) is a key receptor for pheromones involved in regulating reproductive and social behaviors in animals, such as mating between males and females and male-male competition. The aim of this study was to explore variations in circulating androgen and estrogen concentrations in male muskrats in relation to seasonal changes in vomeronasal function. Phylogenetic analysis revealed a total of 212 vomeronasal receptor (VR) genes in muskrats. Previous experimental results demonstrated that among these genes, VN1R41 and VN2R1 act as potential VNO receptors to mediate chemical communication behaviors in muskrats. Therefore, we selected VN1R41 and VN2R1 as representatives of VRs for subsequent experiments. Molecular docking simulations showed that VRs can bind to sex steroid hormones and their sulfated derivatives. Circulating concentrations of testosterone (T), dihydrotestosterone (DHT), and 17β-estradiol (E₂) increased significantly during the breeding season. Type 1 vomeronasal receptors (V1Rs), type 2 vomeronasal receptors (V2Rs), P450arom, androgen receptor (AR), estrogen receptor α (ERα), and estrogen receptor β (ERβ) were detected in both sensory and non-sensory epithelial cells of the VNO in male muskrats. Immunofluorescence assays of VNO also revealed AR and ERα signals in sustentacular cells and sensory cells of the sensory epithelium. The messenger RNA (mRNA) expression levels of VN1R41, AR, ERα, and P450arom in the VNO were significantly higher during the breeding season compared with the nonbreeding season, whereas the mRNA expression levels of VN2R1 and ERβ were lower. Additionally, transcriptomic analysis revealed that differentially expressed genes might be associated with estrogen signaling pathways. These results suggest that androgens and estrogens may regulate vomeronasal function through the expression of AR, ERs, and VRs in the VNO of male muskrats.